Clinical and basic laboratory studies are directed at developing efficient and safe gene transduction strategies for human and hemapoietic cells, including stem and progenitor cells, lymphocytes, and stroma. There are three major ongoing clinical trials of gene transduction for marking purposes only in breast cancer, multiple myeloma, and Gaucher disease. In patients on our service in the Clinical Center, mobilized peripheral blood cells have been shown to be potentially excellent targets for gene transduction, in that they contribute to multilineage long-term engraftment after autologous transplantation. In studies with Gaucher disease, gene marking has been observed at 6 months, even in the absence of myeloablative conditioning. In breast cancer patients treated in the National Cancer Institute, the levels of cells containing marked genes have been very low, possibly related to defective bone marrow function due to previous intensive and repeated courses of hemotherapy. Gene marking and transduction studies have also been performed in rhesus monkeys using autologous transplantation. Ex vivo expansion has been shown to profoundly decrease true stem cell activity. The cytokine called flt 3 ligand and autologous stromal cells may improve retroviral transduction efficiency to levels with clinical utility. The influence of the neomycin resistance gene on transduction has been studied in mouse and the monkey models. The activity of negative regulators of hematopoiesis such as transforming growth factor-beta influences both vector transduction and engraftment by stem cells. Finally, rhesus lymphocytes have been targets for head-to-head comparison of adeno-associated virus and retrovirus, using nonexpressing vectors to eliminate possible immune system effects. Both AAV and retrovirus produce excellent gene transfer levels in vivo in lymphocytes 6 weeks post-infusion.